The present invention concerns therapeutic compositions and methods for the treatment of cancer.
The present invention relates to therapeutic compositions and methods for the treatment or diagnosis of diseases or conditions related to c-fos expression levels, such as cancer. The discussion is not meant to be complete and is provided only for understanding of the invention that follows. This summary is not an admission that any of the work described below is prior art to the claimed invention.
The c-fos proto-oncogene encodes a transcription factor that plays an important regulatory role in the response to mitogenic stimuli (for a review see Angel et al., 1991, Biochem. Biophys. Acta. 1072, 129). Evidence in the literature indicates that c-fos is necessary for expression of many matrix metallo-proteinases (MMPs), including stromelysin 1, stromelysin 2, collagenase 1, matrylisin, 92 kD gelatinase and human macrophage metalloelastase (Sato et al., 1993, Oncogene 8, 395; Gaire et al. 1994 J Biol Chem 269, 2032; Lauricell-Lefebvre et al. 1993 Invasion Metastasis 13, 289; Belaaouajet al. 1995 J Biol Chem 270, 14568). C-fos also regulates the expression of other proteases including urokinase-type plasminogen activator, granzyme B and several cathepsins (Lengyel et al., 1995 Biochim Biophys Acta 1268, 65; Troen et al., 1991 Cell Growth Differ 2, 23; Hadman et al., 1996 Oncogene 12, 135; Rochefort et al., 1995, Ciba Found Symp 191, 254; Wargnier et al. 1995 Proc Natl Acad Sci, USA 92, 6930). Applicant believes that the implications of several of these proteases in tumor metastasis indicates that inhibition of c-fos has the potential to reduce invasive phenotype, as claimed herein. In addition to regulating protease expression, c-fos is necessary for expression of tissue factors, which play an important role in angiogenesis (Felts et al., 1995, Biochemistry 34, 12355; Contrino et al., 1996, Nature Med 2, 209). C-fos is required for expression of the mdr-1 gene (multi-drug resistance), which is thought to contribute to failures in chemotherapy (Scanlon et al., 1994, Proc Natl Acad Sci, USA 91, 11123). C-fos has been shown to play a role in cell proliferation in some cell types (Rijnders et al., 1985, Biochem Biophys Res Comm 132, 548). There is also some suggestion that c-fos may have a role in neuronal injury, degeneration, cell death and/or neoplasms (Schlingensiepen et al., International PCT Publication No. WO 95/02051).
The proto-oncogene c-fos is the cellular homolog of the v-fos gene from FBJ murine osteosarcoma virus. Members of the Fos protein family (c-fos, fosB, fra-1 and fra-2) form heterodimers with members of the jun family (c-jun/AP-1, junB and junD). The heterodimers act as transcriptional activators by binding DNA at AP-1 sites present in a variety of genes, including collagenase, IL-2, adipocyte P2, human metallothionein IIA, transin, and the DNA repair enzymes thymidylate (dTMP) synthase, DNA polymerase .beta., and topoisomerase I. Expression of c-fos is normally tightly regulated at both the RNA and protein level. The kinetics of expression follow the classic pattern of an immediate early gene; mRNA levels peak at 30-45 minutes following mitogenic stimulation and thereafter decline rapidly. The c-fos gene contains an AT-rich mRNA destabilizing sequence in 3' non-coding region, giving the mRNA a half-life of about 12 minutes. The Fos protein has a relatively short half-life (under 2 hours) and negatively regulates transcription of the c-fos gene, contributing to rapid down-regulation (Morgan et al., 1991 Annu Rev Neurosci. 14, 421-451; Ransone et al., 1990, Annu Rev Cell Biol 6, 539).
The connection between fos expression and osteosarcoma was first suggested by the identification of v-fos in murine osteosarcoma virus. Greater than 90% of mice infected with the fos viruses FBJ-MSV and FBR-MSV develop bone tumors. It appears that deregulated expression of the normal c-fos gene can result in similar oncogenic transformation. For example, overexpression of c-fos in tissue culture cells yields a transformed phenotype, and in transgenic mice results in a high frequency of bone and cartilage tumors. The majority of human osteosarcomas (HOS) exhibit significantly elevated c-fos levels (Wu et al. 1990 Oncogene 5, 989). Unlike ras, no specific c-fos mutations have been identified that correlate with oncogenic potential.
Transgenic mice that constitutively express c-fos develop normally until a few weeks after birth, when bone hyperplasia becomes evident (Ruther et al., 1989 Oncogene 4, 861). Approximately 20% develop bone tumors. The level of c-fos expression is at least 10-fold higher in tumor tissue compared to normal tissue. Interestingly, although constitutive expression of c-fos occurs in many tissues, lesions are confined to bone tissue. Thus a secondary tissue-specific event is probably required in addition to elevated c-fos levels to bring about malignant transformation. Fos expression is also associated with cartilage tumor formation when the transgene is expressed during embryogenesis (Wang et al., 1991 EMBO J 10, 2437).
Homozygous c-fos knock-out mice are normal at birth, then begin to exhibit osteopetrosis at about 11 days. This is characterized by severe ossification of the marrow space, shortened bones, and absence of tooth eruption due to obstruction by abnormal amounts of bone. In addition, although possessing normal motor skills, the transgenic animals show behavioral abnormalities including hyperactivity and severely diminished response to external stimuli. This is consistent with reports showing that c-fos plays a pivotal role in the adaptive responses of the nervous system.
Normal bone is constantly being formed and resorbed by the tightly regulated action of osteoblasts and osteoclasts, respectively. This process is controlled in part by parathyroid hormone (PTH) which differentially affects bone mass depending on whether it is present continuously or intermittantly. PTH binds to receptors on osteoblasts and rapidly and transiently induces c-fos expression. PTH-activated osteoblasts then induce c-fos expression in osteoclasts and bone marrow stromal cells. Thus the temporally-regulated expression of c-fos may constitutute an essential downstream event in the normal response to PTH. Either deletion or constitutive overexpression of c-fos in transgenic mice produces abnormal bone morphology, illustrating the requirement for tightly regulated expression of this protein.
Scanlon, International PCT Publication Nos. WO 91/18624 and WO 96/08558; Scanlon et al., 1991, Proc. Natl. Acad. Sci. USA., 88, 10591; and Funato et al., 1992, Advan. Enzyme Regul., 32, 195, report the use of a hammerhead ribozyme to cleave a site within c-fos mRNA.
Scanlon, International PCT Publication No. WO 96/08558, states on page 9-10 that
"[D]rug resistance in mammalian, including human, cancer cells is reversed or ameliorated by the down-regulation of the expression of the Fos/Jun heterocomplex and of AP-responsive genes downstream from Fos/Jun in the transduction pathway. Reversal of MDR phenotype by ribozyme suppression of c-fos oncogene expression illustrates one practical application of the invention."